Plant parasitic nematodes cause serious economic damage to many agricultural crops around the world. These nematodes are microscopic worms and are, in general, obligate parasites of plants. They feed mostly on the roots of host plants; however, several genera are known to parasitize above-ground parts including stem, leaves and flowers as well. Almost all the major plant species are susceptible to infection by species of nematodes (notable exceptions are in the marigolds and asparagus). For example, one of the most important genera of plant nematodes, root-knot nematodes, (Meloidogyne species (spp.)) is capable of parasitizing more than 3,000 species of crop plants. These plants include agronomic crops, vegetables, fruits, flowering trees and shrubs. Nematodes reportedly cause crop loss equivalent to 6 billion dollars in the United States alone and more than 100 billion dollars around the world.
The symptoms due to phytoparasitic nematode injury vary widely depending on the plant host, the nematode species, age of the plant, geographical location, climatic conditions, etc. In general, an overall patchy appearance of plants in a field is considered indicative of nematode injury. More specifically, nematode injury results in galling of the roots (abnormal swelling in the tissues due to rapid multiplication of cells in the cortical region) caused by species of root-knot (Meloidogyne spp.) and cyst (Heterodera spp.) nematodes, lesions (localized, discolored area) caused by lesion nematodes (Pratylenchus spp.), suppression of cell division resulting in "stubby" roots (Trichodorus spp.), growth abnormalities including crinkling or twisting of aboveground parts (Aphelenchoides spp.) and even cell necrosis (death) in some cases. Plant parasitic nematodes may be endoparasitic in nature as in the case of the root-knot and lesion nematodes, or ectoparasitic as in the dagger nematode (Xiphinema spp.) and lance nematode (Hoplolaimus spp.). Nematodes can be vectors of plant viruses and are also known to induce disease complexes by facilitating the entry of other plant pathogenic fungi and bacteria.
Chemical nematocides as soil fumigants or nonfumigants have been in use for many years and represent one of the few feasible processes for countering nematodes. At present, the process involves repeated applications of synthetic chemicals to the ground prior to planting the crop. These chemicals are extremely toxic to organisms other than nematodes and pose a serious threat to the environment. With the renewed emphasis on clean water and air by the United States Environmental Protection Agency (EPA), and the detection of many of these active ingredients or the metabolites thereof in ground water and in several non-target organisms, there has been serious concern about the manufacture and/or use of these chemicals. One of the most effective, economical and widely used nematocides, DBCP (1,2-dibromo-3-chloropropane), was judged to induce male sterility and possible carcinogenesis and was reported in ground water. Another widely used chemical, EDB (ethylene dibromide), was also found in ground water. Yet another very common insecticide-nematocide, aldicarb (2-methyl-2-(methylthio)propionaldehyde-O-(methylcarbamoyl)oxime), was found to be acutely toxic and was found in ground water in several regions of the United States. Carbofuran (2,3-dihydro-2,2-dimethyl-7-benzofuranyl methylcarbamate) and 1,3-D (1,3-dichloropropane), two commonly used nematocides, are under special review by the EPA, because of their avian toxicity and possible carcinogenic effects.
Trichothecenes are a group of closely related sesquiterpenoids that are produced by various species of fungi. Illustrative genera of fungi that produce trichothecenes are Fusarium, Cephalosporium, Myrothecium, Trichothecium, Trichoderma, Cyclindiocarpon, Stachybotrys, Verticimonosporium, Calonectria, and Arthrobotrys.
Several trichothecenes are well-known and commercially available, such as from Sigma Chemical Co., St. Louis, MO. Illustrative trichothecenes include verrucarin A, verrucarol, HT-2 toxin, T-2 toxin, diacetoxyscirpenol, roridin A, acetyl T-2 toxin, neosolaniol, tetracetyl T-2 toxin, T-2 tetraol, T-2 triol, diacetylnivalenol and crotocin.
The structures of the trichothecenes contain a ring system named trichothecane. Godtfredsen et al., Helv. Chim. Acta, 174:1666 (1967). All naturally occurring trichothecene toxins contain an olefinic bond at C-9,10 and most have an epoxy group at C-12,13. Naturally occurring trichothecenes may be classified into five groups (A-E) according to their chemical characteristics.
The trichothecenes of group A possess a hydroxy or an acyloxyl group at C-4 and may have these groups at C-3, C-7, C-8 and C-15, and include HT-2 toxin, T-2 toxin, verrucarol and acetyl T-2 toxin.
The trichothecenes of group B possess a carbonyl group at C-8, and include nivalenol, diacetylnivalenol an trichothecin.
The trichothecenes of group C are macrocyclic trichothecenes and include verrucarin A and roridin A.
The only trichothecene of group D is crotocin which possesses a second epoxide group at C-7,8.
The trichothecenes of group E are trichoverroids which possess either partial or complete carbon chains at C-4 and C-15.
The trichothecenes are stable in the solid state. At extreme pH's, however, the compounds undergo reaction in solution. The esters are saponified by treatment with alkali and the C-12,13-epoxide group is opened by strong mineral acid. Hydrogenation of the C-9,10 double bond results in a slight decrease in toxicity of the compounds, and opening of the C-12,13 epoxide group alters biological activity.
Trichothecenes are known to possess antifungal, antibacterial, antiviral, insecticidal and phytotoxic activity. McDougal et al. (1985) in Progress in the Chemistry of Organic Natural Products (Herz et al., eds. Springer-Verlag, NY) p. 153-219. The trichothecenes have been shown to be toxic to plants at concentrations as low as 10.sup.-7 molar (M), and extremely toxic at concentrations above about 10.sup.-5 M. Cutler (1988) in Biotechnol. for Crop Protect. (Am. Chem. Soc.) p. 50-72.
The nematocidal activity of phytotoxic concentrations of a simple trichothecin (20-500 micrograms (ug) per milliliter (ml)) has been reported by Radzhabova (1971) Dokl. Akad. Nauk. Azerb. SSR, 27:58-60. The trichothecin solution utilized was directly applied to free-living nematodes (Turbatrix aceti) on microscope slides. The trichothecin solutions were prepared in 5% ethanol and a 5% ethanol solution was used as a control. Radzhabova reports that the time period necessary to produce a killing of 50% of the nematodes tested (LD.sub.50) is significantly reduced as the trichothecin concentration increases. A solution having 500 ug/ml trichothecin produced an LD.sub.50 about 4-fold lower than that for a 62.5 ug/ml trichothecin solution and an LD.sub.50 approximately one-half that produced by a 250 ug/ml trichothecin solution in the assay. Since it is known that many species of nematodes are susceptible to ethanol, this article does not teach that trichothecenes are nematocidal and in no way discloses the nematostatic activity of trichothecenes that is part of the present invention.